The invention relates to a force measurement of the sensitivity threshold of animals or humans, and in particular the sensitivity or nociciptive (pain) threshold of rats and mice as is needed for medical research, such as to measure analgesic effects.
The measurement of mechanical sensitivity, or nociceptive (pain) threshold has long been used in many areas of medical research, in particular as a measure of analgesic effect. The hypothesis is that, after a force (or pressure) threshold figure has been established without treatment, the administration of an analgesic will elevate that threshold.
In humans, and in larger animal species, so called mechanical algometers, palpometers, dolorimeters or aesthesiometers are used. These are essentially hand-held force gauges, with a tip of suitable cross-sectional area and profile for the species and test site. The tip is pushed with a progressively increasing force into the surface tissue of the subject until a response is elicited. This is assumed to be the nociceptive threshold. The response may be vocal or behavioral. Ethics demand that the stimulus is removed rapidly at the threshold.
Such methods become difficult to implement at low forces, as are required for small animals such as rats and mice. The force transducer becomes unwieldy compared to the size of the animal and, if the force is applied by hand, overly sensitive to the inevitable slight hand tremor of the operator.
Historically, “von Frey filaments” have been used to overcome these problems in small animals [1, 2]. Von Frey filaments are sold by Ugo Basile for example [3]. Von Frey filaments are thin filaments of a flexible material with a high elastic strain range, supplied as a set of differing diameters but of the same length. As the diameter increases, so does the compressive force required for buckling. The diameters of the filaments are arranged so that, when a compressive force is applied along their axis, the buckling forces are in useful increments of the total force range to be investigated. The range of von Frey filaments or “hairs” typically runs from 0.008 g to 300 g. The highest diameter is about 1.3 mm and the lowest less than 0.1 mm. (In practice, presumably because of manufacturing restrictions, these force increments are very un-equal.) Thus the successive application of progressively thicker filaments to the test site on the subject will provide a force which increases in a number of un-equal steps.
Typically, for the measurement of mechanical thresholds on rats and mice, the ventral surface of the paw is used for testing, with the animal allowed to move in 2 axes in a cage whose base is formed of an open grid. Von Frey filaments are advanced upwards towards the pad of the paw by the tester, through the grid. Typically, the behavioral response is for the animal to lift its foot.
Considerable judgment is required by the tester as, at some point during the successive application of a series of von Frey filaments of progressively increasing diameter, the force will become sufficient to simply lift the foot, rather than the animal withdrawing it voluntarily. This is not a valid test.
In summary, von Frey filaments have a number of drawbacks:
1. The test site is subjected to a sequence of tests before a result is reached. If the subject reacts to the first filament, then only one test is made and the result is limited to having established that the threshold force is the force associated with the first filament or less. If reaction occurs later during a second or further test, the result is still only within a band of forces defined by the buckling force separation between adjacent filament sizes used in the test. This is time consuming and undoubtedly has implications for a behavioral response. Previous stimuli may also change the perception of subsequent ones.
2. As the filaments are, of necessity, of different diameters (in order to achieve a progressively increasing force), the area of tissue on which they act also changes (each filament is cleaved across its end to achieve a nominally flat disc of contact). As the area of tissue changes so does the pressure beneath the surface (pressure=force/area), and potentially also the number of nociceptors that are triggered by the stimulus.
3. The buckling load for each filament, while repeatable when the filament is new and undamaged, reduces substantially as soon as the filament takes a set or curve, through use, misuse or damage.
4. The contact area changes as the filament buckles, from a flat disc to a curved line edge contact. This changes the feel of the filament at the threshold and may have implications for the animal's response to the next test.
5. The use of a number of successive tests requires the test subject to remain stationary. Healthy rats and mice are generally disinclined to do this. The experimenter, viewing a square or rectangular cage from the side, must therefore follow the rodent while attempting to push the filaments through the gaps in the grid without touching the grid (the friction from which would invalidate the test).
6. Since the transducing element is stiff up to the point at which it buckles, hand tremor from the operator is transmitted as a variation in force during the force ramp, although the maximum force cannot exceed the rated force for that filament.
There have been, in recent years, a number of attempts to produce alternatives to the traditional von Frey systems, for example:                Electronic von Frey Anethesiometer, model 2390        IITC Inc Life Science        23924 Victory Blvd        Woodland Hills        Calif. 91367        Electronic von Frey, model BSEVF3        Harvard Apparatus        Fircroft Way        Edenbridge        Kent        TN8 6HE        United Kingdom        37450 Dynamic Plantar Aesthesiometer [4]        Ugo Basile North America Inc        414 West Main Street        Collegeville        Pa. 19426        Electronic Von Frey Sensebox [5]        Somedic AB        Box 194        SE-242 22 Horby        Sweden        
These solutions are, in one form or another, low capacity force gauges with a single thin probe to apply the force. These systems are generally difficult to use, for two reasons:
Firstly, the force producing and sensing mechanism is influenced greatly by any hand tremor of the operator. This is because the system is fundamentally much stiffer than a von Frey filament over the entire measurement range: any small movement results in a relatively large change of force. This may be seen by attempting to push down with a small but constant force on a set of digital kitchen scales (where the deflection of the force transducer within will be millimeters or fractions of millimeters for the entire force range); the reading will be seen to fluctuate considerably due to hand tremor. A von Frey filament, in contrast, is only stiff up to the point at which it buckles; at that point it becomes a very soft spring where the force applied is relatively insensitive to hand tremor.
Secondly, for laboratory animals, the time window available for a test is small (perhaps 3-5 seconds) before the animal moves to a different location within the cage. Some of the systems available require the operator to follow a force/time ramp on a computer screen while applying the force. This is impractical while looking at the mouse's foot and unrealistic in the time frame available.
Attempts have also been made to automate the force application process by means of a motor driven probe which advances up through the grid to the foot from a movable platform. This platform must still be manually positioned relative to the foot in X and Y by the operator (using a mirror), and requires the animal to remain motionless for the time of the test (5 seconds or more).
The problems outlined above apply to both rats and mice. They are however greater for mice, due to their smaller size and correspondingly lower threshold force. As an example, mechanical thresholds for mice are in the order of 1-4 grams force, and up to 50 grams force or more for rats. As laboratory mice make up 90% of the animal laboratory rodent population, and therefore a similar proportion of all nociceptive testing, there is scope for improvement of the current approaches.